Single-stage autotrophic removal of thiocyanate and nitrogen from high-strength thiocyanate wastewater: start-up, performance, and metabolic mechanisms.

Abstract

A single-stage partial nitrification and thiocyanate-driven denitrification (SPN-TDN) process offers a promising and resource-efficient route for simultaneous thiocyanate and nitrogen removal from high-strength thiocyanate (SCN-) wastewater. However, reliable start-up remains challenging because the establishment of this process requires coordination under contrasting oxygen and substrate conditions. Here, a sequencing batch reactor was operated for 384 cycles using a staged enrichment-optimization strategy to investigate how the synergistic process became established under increasing SCN- loading. During enrichment, SCN- removal remained above 98 %, partial nitrification was established, but total nitrogen (TN) removal stayed limited. In the optimization phase, TN removal progressively increased to 40 %, accompanied by biogenic elemental sulfur accumulation. Ex-situ batch tests confirmed SCN--driven autotrophic denitrification, achieving 49 % TN removal with a nitrite/SCN- consumption ratio of 0.38. Community profiling showed that phase I established the dominant Thiobacillus (0.2 % to 23.0 %) and Nitrosomonas (0.99 % to 2.46 %), with the normalized stochasticity ratio indicating a greater deterministic contribution at both genus and species levels. Phase II largely retained this genus-level framework while exhibiting taxonomic-scale-dependent assembly and lineage-level turnover. Genome-resolved metagenomics further showed increased representation of cyanate-pathway lineages, a corresponding decline in the carbonyl sulfide-pathway lineages, and lineage-level turnover within ammonia-oxidizing bacteria. Collectively, successful process establishment involved multiple functional pathways during enrichment and subsequent pathway- and lineage-level reorganization under intensified constraints. These findings provide a mechanistic basis for understanding and further optimizing staged start-up of single-stage SPN-TDN for high-strength SCN- wastewater treatment.